Parkinson's disease (PD) is a common neurological condition that progresses from subtle loss of muscular coordination to severe physical and mental disability and eventual death. There is no cure and no treatments to slow the disease progression. Currently there are ~1 million PD sufferers in the US, which costs about $25 billion USD per year. These numbers are expected to triple by 2050 as the population of the elderly is increasing rapidly. Thus, effective disease-modifying treatments are desperately needed. To contribute to the drug development effort, the goal of this project is to determine the structure o -synuclein (-syn) to gain insights into its function and its mechanisms in the pathogenesis of Parkinson's disease, which are critical for rational drug design. The aggregation of -syn leads to appearance of so-called Lewy bodies (LBs) as a defining pathological hallmark of Parkinson's disease. Similar aggregates of -syn are also implicated in other neurodegenerative diseases, including multiple system atrophy (MSA), dementia with Lewy bodies (DLB), and others collectively known as synucleinopathies. -Syn is highly abundant in the brain and it appears to be important for learning and memory; however, its precise function is still unknown. It has no known enzymatic activity, thus its unknown biological function depends on its physical structure; moreover, it causes disease via a gain-of-toxic function that is associated with its structural transformation into highly ordered fibrils. Thus, detailed structural information of -syn is crucil for understanding its function and to gain insights into its transformation into a disease-causing entity. However, such structural details are currently unavailable mainly because, until recently, -syn lacked a persistent structure in solution and was thought to be a natively unfolded protein, which is not amenable for structure determination. However, we have recently isolated an ordered tetrameric form of -syn in solution. We did so by developing an expression and purification procedure aimed at preserving its native structure throughout the purification process. At about the same time, Dennis Selkoe's group at Harvard Medical School independently demonstrated that -syn isolated from rat brains and living human cells are also natively folded and tetrameric with striking resemblance to our recombinant -syn. Moreover, we further demonstrated that our -syn preparation was resistant to aggregation, did not compromise liposome membranes, and non-toxic to cells, thus strongly supporting that the ordered tetrameric -syn is physiologically relevant. Now that we have an ordered form of -syn that is amenable to traditional biophysical tools, the specific aims of this proposal are; 1) To determine the atomic structure of -syn; and 2) To determine the dynamics of its quaternary structure. We will do so by using a combination of X-ray crystallographic methods, nuclear magnetic resonance (NMR) methods, and single-molecular fluorescence methods. PUBLIC HEALTH RELEVANCE: This study proposes to determine the molecular structure of -synuclein and to determine the molecular kinetics of its tetrameric structure assembly, which will provide the molecular basis and a framework for rational drug design to combat Parkinson's disease.